Detecting minimal residual disease in neuroblastoma: the superiority of a panel of real-time quantitative PCR markers.

BACKGROUND: PCR-based detection of minimal residual disease (MRD) in neuroblastoma (NB) patients can be used for initial staging and monitoring therapy response in bone marrow (BM) and peripheral blood (PB). PHOX2B has been identified as a sensitive and specific MRD marker; however, its expression varies between tumors. Therefore, a panel of markers could increase sensitivity. METHODS: To identify additional MRD markers for NB, we selected genes by comparing SAGE (serial analysis of gene expression) libraries of healthy and NB tissues followed by extensive real-time quantitative PCR (RQ-PCR) testing in samples of tumors (n = 56), control BM (n = 51), PB (n = 37), and cell subsets. The additional value of a panel was determined in 222 NB samples from 82 Dutch stage 4 NB patients (54 diagnosis BM samples, 143 BM samples during/after treatment, and 25 PB samples). RESULTS: We identified 2 panels of specific RQ-PCR markers for MRD detection in NB patients: 1 for analysis of BM samples (PHOX2B, TH, DDC, CHRNA3, and GAP43) and 1 for analysis of PB samples (PHOX2B, TH, DDC, DBH, and CHRNA3). These markers all showed high expression in NB tumors and no or low expression in control BM or PB samples. In patients' samples, the PHOX2B marker detected most positive samples. In PB samples, however, 3 of 7 PHOX2B-negative samples were positive for 1 or more markers, and in BM examinations during treatment, 7% (6 of 86) of the PHOX2B-negative samples were positive for another marker. CONCLUSIONS: Because of differences in the sensitivities of the markers in BM and PB, we advise the use of 2 different panels to detect MRD in these compartments.

PHOX2B is a novel and specific marker for minimal residual disease testing in neuroblastoma.

PURPOSE: Polymerase chain reaction (PCR)-based detection of minimal residual disease (MRD) in neuroblastoma can be used to monitor therapy response and to evaluate stem cell harvests. Commonly used PCR markers, tyrosine hydroxylase (TH) and GD2 synthase, have expression in normal tissues, thus limiting MRD detection. To identify a more specific MRD marker, we tested PHOX2B. PATIENTS AND METHODS: To determine PHOX2B, TH, and GD2 synthase expression in normal tissues, it was measured by real-time quantitative PCR in samples of normal bone marrow (BM; n = 51), peripheral blood (PB; n = 37), and peripheral-blood stem cells (PBSCs; n = 24). Then, 289 samples of 101 Dutch patients and 47 samples of 43 German patients were tested for PHOX2B and TH; these samples included 52 tumor, 214 BM, 32 BM, and 38 PBSC harvests. Of the 214 BM samples, 167 were compared with cytology, and 47 BM samples were compared with immunocytology (IC). RESULTS: In contrast to TH and GD2 synthase, PHOX2B was not expressed in any of the normal samples. In patient samples, PHOX2B was detected in 32% cytology-negative and in 14% IC-negative samples and in 94% of cytology-positive and in 90% of IC-positive BM samples. Overall, PHOX2B was positive in 43% compared with 31% for TH. In 24% of all samples, TH expression was inconclusive, which is similar to expression found in normal tissues. In 42% of these samples, PHOX2B expression was positive. CONCLUSION: PHOX2B is superior to TH and GD2 synthase in specificity and sensitivity for MRD detection of neuroblastoma by using real-time quantitative PCR. We propose to include PHOX2B in additional prospective MRD studies in neuroblastoma alongside TH and other MRD markers.

An exclusive case of juvenile myelomonocytic leukemia in association with Kikuchi's disease and hemophagocytic lymphohistiocytosis and a review of the literature.

We present a case of juvenile myelomonocytic leukemia (JMML) accompanied by immune-mediated hemophagocytic lymphohistiocytosis (HLH) and Kikuchi's disease, both as a paraneoplastic phenomenon. As this combination, to the best of our knowledge, has not been described before, consensus on preferable treatment is lacking. Our patient was treated with prednisolone according to the few described cases of HLH and Kikuchi's disease in non-JMML patients, resulting in disappearance of the clinical symptoms.